E. coli-derived human Sonic Hedgehog/Shh protein Cys24-Gly197 (Cys24Ile-Ile), with an N-terminal Met Produced using non-animal reagents in an animal-free laboratory.
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain.
<0.01 EU per 1 μg of the protein by the LAL method.
N-terminal sequence Analysis
Predicted Molecular Mass
22 kDa, reducing conditions
Measured by its ability to induce alkaline phosphatase production by C3H10T1/2 mouse embryonic fibroblast cells. Nakamura, T. et al. (1997) Biochem. Biophys. Res. Commun. 237:465. The ED 50 for this effect is 0.1-0.4 µg/mL.
Recombinant Human Sonic Hedgehog/Shh (C24II) Animal-Free, CF Scientific Data Examples
Recombinant Human Sonic Hedgehog/SHH (C23IL) Animal-Free Bioactivty.
Animal-Free™ Recombinant Human Sonic Hedgehog/Shh (Catalog # AFL1845) induces alkaline phosphatase production by the C3H10T1/2 mouse embryonic fibroblast cell line. The ED50 for this effect is 0.1-0.4 μg/mL.
Recombinant Human Sonic Hedgehog/Shh (C24II), Animal Free Protein SDS-PAGE.
1 μg/lane of Animal-Free™ Recombinant Human Sonic Hedgehog/Shh (C24II) Protein (Catalog # AFL1845) was resolved with SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by silver staining, showing bands at 22 kDa.
Formulation, Preparation and Storage
What does CF mean?
CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our
Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant
protein to be stored at a more dilute concentration.
The carrier free version does not contain BSA.
What formulation is right for me?
In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or
as an ELISA standard.
In contrast, the carrier free protein is recommended for applications, in which the presence of BSA
Lyophilized from a 0.2 μm filtered solution in PBS and NaCl.
Reconstitute at 100-200 μg/mL in PBS, and allow up to 24 hours at 2 to 8 degreesC for complete reconstitution.
The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage:
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
12 months from date of receipt, -20 to -70 degreesC as supplied. 1 month, 2 to 8 degreesC under sterile conditions after reconstitution. 3 months, -20 to -70 degreesC under sterile conditions after reconstitution.
Background: Sonic Hedgehog/Shh
Sonic Hedgehog (Shh) is expressed in embryonic tissues that are critical for the patterning of the developing central nervous system, somite, and limb. It is also involved in whisker, hair, foregut, tooth, and bone development. Shh regulates neural and hematopoietic stem cell fate and is important for thymocyte differentiation and proliferation as well as T cell determination. In adult tissue Shh is associated with cancer development and tissue remodeling following injury (1-3). Human Shh encodes a 462 amino acid (aa) precursor protein that is autocatalytically processed to yield a non-glycosylated 19 kDa N-terminal fragment (Shh-N) and a glycosylated 25 kDa C-terminal protein (Shh-C) (4). Shh-C, which is responsible for the intramolecular processing of Shh, is rapidly degraded following Shh proteolysis (5). Shh-N is highly conserved, sharing >98% aa identity between mouse, human, rat, canine, porcine, and chicken Shh-N. Shh-N can be palmitoylated at its N-terminal cysteine and modified by cholesterol addition at its C-terminus (6). These modifications contribute to the membrane tethering of Shh as well as its assembly into various sized multimers (6-9). Lipid modification and multimerization greatly increase Shh-N receptor binding affinity and signaling potency (5, 6, 8, 9). Monomeric and multimeric Shh can be released from the plasma membrane by the cooperative action of DISP1, SCUBE2, and TACE/ADAM17 (10-12). Modifications also extend the effective range of Shh functionality and are required for the development of protein gradients important in tissue morphogenesis (9, 13). Canonical signaling of Shh is mediated by a multicomponent receptor complex that includes Patched (PTCH1, PTCH2) and Smoothened (SMO) (14). The binding of Shh to PTCH releases the basal repression of SMO by PTCH. Shh activity can also be regulated through interactions with heparin, glypicans, and membrane-associated Hip (hedgehog interacting protein) (13, 15, 16).
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Aviles, E.C. et al. (2013) Front. Cell. Neurosci. 7:86.
Xie, J. et al. (2013) OncoTargets Ther. 6:1425.
Marigo, V. et al. (1995) Genomics 28:44.
Zeng, X. et al. (2001) Nature 411:716.
Feng, J. et al. (2004) Development 131:4357.
Goetz, J.A. et al. (2006) J. Biol. Chem. 281:4087.
Pepinsky, R.B. et al. (1998) J. Biol. Chem. 273:14037.
Chen, M.-H. et al. (2004) Genes Dev. 18:641.
Etheridge, L.A. et al. (2010) Development 137:133.
Jakobs, P. et al. (2014) J. Cell Sci. 127:1726.
Dierker, T. et al. (2009) J. Biol. Chem. 284:8013.
Lewis, P.M. et al. (2001) Cell 105:599.
Carpenter, D. et al. (1998) Proc. Natl. Acad. Sci. USA 95:13630.
Filmus, J. and M. Capurro (2014) Matrix Biol. 35:248.
Chuang, P.-T. and A.P. McMahon (1999) Nature 397:617.
Entrez Gene IDs
6469 (Human); 20423 (Mouse)
HHG-1, HHG1, HLP3, HPE3, MCOPCB5, MCOPCB5sonic hedgehog (Drosophila) homolog, SMMCI, SMMCIsonic hedgehog homolog (Drosophila), Shh, ShhNC, Sonic Hedgehog, TPT, TPTPS, sonic hedgehog homolog, sonic hedgehog protein
Animal-Free Manufacturing Conditions Our dedicated controlled-access animal-free laboratories ensure that at no point in production are the products exposed to potential contamination by animal components or byproducts. Every stage of manufacturing is conducted in compliance with R&D Systems' stringent Standard Operating Procedures (SOPs). Production and purification procedures use equipment and media that are confirmed animal-free.
All molecular biology procedures use animal-free media and dedicated labware.
Dedicated fermentors are utilized in committed animal-free areas.
Protein purification columns are animal-free.
Bulk proteins are filtered using animal-free filters.
Purified proteins are stored in animal-free containers in a dedicated cold storage room.
Low Endotoxin Level.
No impairment of biological activity.
High quality product obtained under stringent conditions.
Which Brands are Currently Available on bio-techne.com? R&D Systems, Tocris Bioscience and ProteinSimple branded products are available to purchase through bio‑techne.com. ProteinSimple branded instruments and Simple Plex Assays are available to quote. ProteinSimple branded instrument consumables and ACD branded products will be available on bio-techne.com in the near future. Novus Biologicals branded products are not currently available on bio‑techne.com and can be found at novusbio.com.
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